Reduced Frizzled-Related Protein B Expression Is Associated With Wnt/β-Catenin Signaling Activation and Calcification in Degenerative Menisci

Abstract

Meniscal calcification is increasingly recognized as a pathological feature associated with the progression of osteoarthritis (OA). However, the molecular mechanisms underlying this process remain poorly understood. This study aimed to investigate the involvement of Wnt/β-catenin signaling and its extracellular antagonist frizzled-related protein B (FRZB) in pathological calcification of degenerative human menisci. Human meniscal specimens were obtained from patients with knee OA and non-OA controls and analyzed histologically and immunohistochemically for calcification and expression of hypertrophic/calcification-associated markers, β-catenin, and FRZB. An in vitro calcification model using primary human meniscal cells was established, and the role of FRZB was examined by supplementation with recombinant FRZB or shRNA-mediated FRZB knockdown. Calcification was assessed with Alizarin Red S staining, and expression of Wnt/β-catenin signaling-related and hypertrophic/calcification-associated genes was analyzed using quantitative PCR and western blotting. Degenerative menisci exhibited prominent calcium deposition accompanied by increased expressions of RUNX2, collagen type X, and MMP-13. Calcified menisci showed enhanced β-catenin expression and reduced FRZB expression compared with non-calcified menisci. In an in vitro model, meniscal cell calcification was accompanied by downregulation of FRZB and activation of Wnt/β-catenin signaling, along with upregulation of RUNX2, collagen type X, and MMP-13. Recombinant FRZB attenuated Wnt/β-catenin signaling activity and reduced calcification, whereas FRZB knockdown enhanced Wnt/β-catenin signaling and increased calcium deposition. Reduced FRZB expression was associated with enhanced Wnt/β-catenin signaling activity and increased calcification in degenerative menisci. The findings suggest a potential inhibitory role for FRZB in the context of Wnt/β-catenin signaling associated meniscal calcification.